The experiments outlined in this proposal will examine the medical and health consequences of cocaine abuse, which has been identified as NIDA as a serious problem in this country. Since 1998 the use of cocaine in the US has risen, in part due to the low cost and ample supply. Paralleling this trend has been an increase in the rate of emergency room admissions related to cocaine use and cocaine-related deaths. The exact causes of the cocaine-related fatalities in most cases are not known but many are believed to result from cardiac arrhythmias, the leading cause of death associated with cocaine abuse. In humans, cocaine is known to increase heart rate and blood pressure, effects that are primarily attributable to the sympathomimetic actions of this drug. Cocaine also produces significant changes in cardiac electrophysiology, decreasing the conduction and slowing the repolarization following an action potential. These electrical disturbances are believed to result from a direct anesthetic effect of cocaine on cardiac ion channels. In this proposal, we will investigate the cocaine sensitivity of sodium and potassium channels important for initiating and terminating the cardiac action potential. Inhibiting these channels disrupts the coordinated electrical activity of the heart, a well-known risk factor for the generation of ventricular arrhythmias. A combination of electrophysiology and molecular biology will be used to investigate the mechanisms of inhibition of these channels and to identity sites important for cocaine binding. The human cardiac sodium channel (hH1) and the human ether-a-g-go delayed rectifier potassium channel (HERG) will be expressed in mammalian cells and the resulting currents recorded using patch clamp techniques. Our data indicate that cocaine binding to these channels is state-dependent, with cocaine preferentially binding to the open and inactivated channels. Inactivation-deficient mutants of hH1 and HERG will be constructed to investigate the contribution of inactivation gating to cocaine binding and to facilitate the measurement of open-channel block. Additional mutants with the sixth transmembrane spanning segments, the putative anesthetic binding sites of these channels, will also be investigated. The proposed studies ill provide insight into the mechanisms underlying the cocaine-induced changes in cardiac electrophysiology and the cardiotoxic effects of this drug.